Towards engineering a multi-cell lineage multi-organism intestine

This proposal is aligned to two of the NC3Rs objectives, namely the Replacement and Reduction of animals in research. The establishment of a system whereby the gut can be recapitulated in vitro will have serious impacts upon the number of animals required for use in research. The intestine fulfils critical roles for life. Its physiology and homeostasis requires complex cellular and molecular interactions. Gene-targeted mice and infection models have provided important insights into enteric physiology, but the complexity of the in vivo system hinders detailed study of cellular and molecular components and requires the use of large cohorts of model animals to observe statistical differences. We will adapt organoid cultures (a rudimentary gut) to incorporate cells critical to intestinal homeostasis. This enables the study of interactions of immune cells with gut-epithelial cells, important in tissue turnover and repair, in great detail and with minimal use of model animals and without regulated procedures. To meet the challenge of complexity and future use of the system, we will use interdisciplinary approaches at the interface of immunology, microbiology and bioengineering. This will form the basis for the development of highly organized intestines with sealed enteric luminal and basolateral sides, ensuring the full development of a 3D multi-organism artificial intestine. This enables the study of the gut in steady state and in varying contexts of growth factors, dietary compounds, micro-organisms and immune cells. The challenge to create sealed luminal and basolatheral sides does not preclude to study the interactions of the epithelial cells with micro-organisms. We will initiate this by using conserved micro-organismal products and test their effect on cells of the intestine in the absence and presence of immune cell subsets. We will also test the suitability of the system in providing the correct conditions for the full lifecycle of intestinal parasite species which critically rely on host factors in the intestine.